The present invention relates to novel cytotoxic agents with enhanced water solubility and their therapeutic uses. More specifically, the invention relates to novel cytotoxic agents that are taxanes which comprise both a polyethylene glycol moiety that enhances water solubility and a means of chemical linkage to a cell binding agent. These taxanes can be chemically linked to cell binding agents to provide therapeutics that are delivered to specific cell populations in a targeted manner.
Many reports have appeared which are directed to the tageting of tumor cells with monoclonal antibody-drug conjugates (Sela et al, in Immunoconjugates, pp. 189-216 (C. Vogel, ed. 1987); Ghose et al, in Targeted Drugs, pp. 1-22 (E. Goldberg, ed. 1983); Diener et al, in Antibody Mediated Delivery Systems, pp. 1-23 (J. Rodwell, ed. 1988); Pietersz et al, in Antibody Mediated Delivery Systems, pp. 25-53 (J. Rodwell, ed. 1988); Bumol et al, in Antibody Mediated Delivery Systems, pp. 55-79 (J. Rodwell, ed. 1988), G. A. Pietersz and K. Krauer, 2 J. Drug Targeting, 183-215 (1994), R. V. J. Chari, 31 Adv. Drug Delivery Revs., 89-104 (1998); W. A. Blattler and R. V. J. Chari, in Anticancer Agents, Frontiers in Cancer Chemotherapy, 317-338, ACS Symposium Series 796, I. Ojima et al eds, American Chemical Society 2001). Cytotoxic drugs such as methotrexate, daunorubicin, doxorubicin, vincristine, vinblastine, melphalan, mitomycin C, chlorambucil, calicheamicin and maytansinoids have been conjugated to a variety of murine monoclonal antibodies. In some cases, the drug molecules were linked to the antibody molecules through an intermediary carrier molecule such as serum albumin (Garnett et al, 46 Cancer Res. 2407-2412 (1986); Ohkawa et al, 23 Cancer Immunol. Immunother. 81-86 (1986); Endo et al, 47 Cancer Res. 1076-1080 (1980)), dextran (Hurwitz et al, 2 Appl. Biochem. 25-35 (1980); Manabi et al, 34 Biochem. Pharmacol. 289-291 (1985); Dillman et al, 46 Cancer Res. 4886-4891 (1986); Shoval et al, 85 Proc. Natl. Acad. Sci. U.S.A. 8276-8280 (1988)), or polyglutamic acid (Tsukada et al, 73 J. Natl. Canc. Inst. 721-729 (1984); Kato et al, 27 J. Med. Chem. 1602-1607 (1984); Tsukada et al, 52 Br. J. Cancer 111-116 (1985).
A wide array of linkers is now available for the preparation of such immunoconjugates, including both cleavable and non-cleavable linkers. In vitro cytotoxicity tests, however, have revealed that antibody-drug conjugates rarely achieve the same cytotoxic potency as the free unconjugated drugs. This suggested that mechanisms by which drug molecules are released from conjugated antibodies are very inefficient. Early work in the area of immunotoxin conjugates showed that conjugates formed via disulfide bridges between monoclonal antibodies and catalytically active protein toxins were more cytotoxic than conjugates containing other linkers. See, Lambert et al, 260 J. Biol. Chem. 12035-12041 (1985); Lambert et al, in Immunotoxins 175-209 (A. Frankel, ed. 1988); Ghetie et al, 48 Cancer Res. 2610-2617 (1988). This improved cytotoxicity was attributed to the high intracellular concentration of reduced glutathione contributing to the efficient cleavage of the disulfide bond between the antibody molecule and the toxin. Maytansinoids and calicheamicin are the first examples of highly cytotoxic drugs that have been linked to monoclonal antibodies via disulfide bonds. Antibody conjugates of these drugs have been shown to possess high potency in vitro and exceptional antitumor activity in human tumor xenograft models in mice (R. V, J. Chari et al., 52 Cancer Res., 127-131 (1992), C. Liu et al., 93, Proc. Natl. Acad. Sci., 8618-8623 (1996), L. M. Hinman et al., 53, Cancer Res., 3536-3542 (1993), P. R. Hamann et al, 13, BioConjugate Chem., 40-46 (2002)).
One reason for the lack of disulfide linked antibody-drug conjugates is the unavailability of cytotoxic drugs possessing a sulfur atom containing moiety that can be readily used to link the drug to an antibody via a disulfide bridge. Furthermore, chemical modification of existing drugs is difficult without diminishing their cytotoxic potential.
In spite of the above described difficulties, useful cytotoxic agents comprising cell binding moieties and the group of cytotoxic drugs known as maytansinoids have been reported (U.S. Pat. No. 5,208,020, U.S. Pat. No. 5,416,064, and R. V. J. Chari, 31 Advanced Drug Delivery Reviews 89-104 (1998)). Similarly, useful cytotoxic agents comprising cell binding moieties and analogues and derivatives of the potent antitumor antibotic CC-1065 have also been reported (U.S. Pat. No. 5,475,092 and U.S. Pat. No. 5,585,499).
Paclitaxel (Taxol(copyright)), a cytotoxic natural product, and docetaxel (Taxotere(copyright)), a semi-synthetic derivative (See FIG. 1), are widely used in the treatment of cancer. These compounds belong to the family of compounds called taxanes. Taxanes are mitotic spindle poisons that inhibit the depolymerization of tubulin, resulting in an increase in the rate of microtubule assembly and cell death. While docetaxel and paclitaxel are useful agents in the treatment of cancer, their antitumor activity is limited because of their non-specific toxicity towards normal cells.
Further, compounds like paclitaxel and docetaxel themselves are not sufficiently potent to be used in conjugates of cell binding agents. Recently, a few new taxanes with greater potency than either docetaxel or paclitaxel have been prepared (FIG. 1). In addition, these taxanes bear a suitable functionality that allows linkage via a cleavable bond to cell binding agents (U.S. Pat. Nos. 6,340,701 and 6,372,738). However, these taxanes display poor aqueous solubility. Thus linkage to cell binding agents, which are typically only soluble in water, necessitates the use of a high percentage of an organic co-solvent which could lead to damage of the cell binding agent. Thus, conjugation reactions with cell-binding agents currently have to be performed in extremely dilute aqueous solutions.
One approach commonly used to enhance the aqueous solubility of poorly soluble drugs such as paclitaxel is to convert them into prodrugs by incorporating a polyethylene glycol spacer of varying chain lengths, in a process often called PEGylation. These prodrugs are inactive or poorly active in vitro, and have to rely on in vivo enzymatic cleavage of the polyethylene glycol group to be activated. Such in vivo cleavage mechanisms are inefficient resulting in poor conversion into active drug. In addition, these PEGylated-taxanes do not have a linking group that allows for conjugation to cell binding agents (U.S. Pat. Nos. 5,614,549; 5,648,506; 5,880,131; 5,824,701; R. B. Greenwald et al., 60, J. Org. Chem., 331-336 (1995), R. B. Greenwald et al., 39, J. Med. Chem., 424-431 (1996), A. E. Matthew et al., 35, J. Med. Chem., 145-151 (1992)).
Accordingly, a method of providing taxanes that contain a polyethylene glycol moiety that confers enhanced aqueous solubility, while preserving cytotoxic potency, without the need for additional in vivo activation is required. These PEGylated taxanes must also possess a linking group that allows for linkage with cell binding agents. Thus, a method of using these taxanes for treating diseases wherein their side effects are reduced without compromising their cytotoxicity is greatly needed.
One object of the present invention is to provide novel taxanes that incorporate a polyethylene glycol moiety that confers enhanced aqueous solubility.
Another object of the present invention is to provide polyethylene glycol-containing taxanes that are highly cytotoxic in vitro and that can still be effectively used in the treatment of many diseases.
These and other objects have been achieved by providing a taxane comprising a polyethylene glycol-containing linking group at C-7 or C-10, the linking group being capable of linking the taxanes to a cell binding agent or other chemical moiety.
The invention also provides a cytotoxic agent comprising one or more taxanes linked to a cell binding agent through a polyethylene glycol-containing linking group at C-7 or C-10 of at least one of the taxanes.
The present invention also provides a therapeutic composition comprising an effective amount of a cytotoxic agent comprising one or more taxanes linked to a cell binding agent through a polyethylene glycol-containing linking group at C-7 or C-10 of at least one of the taxanes; and (B) A pharmaceutically acceptable carrier, diluent, or excipient.
The present invention also provides a method of killing selected cell populations comprising contacting the target cells or tissue containing target cells with a cytotoxic amount of the above-described cytotoxic agent.